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Dissimilatory Nitrate Reductase from Bradyrhizobium sp. (Lupinus): Subcellular Location, Catalytic Properties, and Characterization of the Active Enzyme Forms

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Abstract

Subcellular location, chlorate specificity, and sensitivity to micromolar concentrations of azide suggest that most of the anaerobically induced nitrate reductase (NR) activity in Bradyrhizobium sp. (Lupinus) could be ascribed to the membrane type of bacterial dissimilatory NRs. Two active complexes of the enzyme, NRI of 140 kDa and NRII of 190 kDa, were detected in membranes of the nitrate-respiring USDA strain 3045. Both enzyme forms were purified to homogeneity. Obtained specific antibodies showed that these native species were immunologically closely related and composed of largely similar 126-kDa, 65-kDa, and 25-kDa subunits. The finding that NRI and NRII share common epitopes suggests that they may not be different species, but rather two forms of the same enzyme.

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Literature Cited

  1. Alikulov ZA, Burikhanov SS, Sergeev NS, Lvov NP, Kretovich VL (1980) Nitratreduktaza bakteroidov klubienkov lupina. Prikl Biochim Mikrob (Transl) 16:511–516

    CAS  Google Scholar 

  2. Bedzyk L, Wang T, Ye RW (1999) The periplasmic nitrate reductase in Pseudomonas sp. strain G-179 catalyzes the first step of denitrification. J Bacteriol 181:2802–2806

    CAS  PubMed  Google Scholar 

  3. Bell LC, Richardson DJ, Ferguson SJ (1990) Periplasmic and membrane-bound respiratory nitrate reductases in Thiosphaera pantotropha. FEBS Lett 265:85–87

    Article  CAS  PubMed  Google Scholar 

  4. Berks BC, Ferguson SJ, Moir JW, Richardson DJ (1995) Enzymes and associated electron transport systems that catalyse the respiratory reduction of nitrogen oxides and oxyanions. Biochim Biophys Acta 1232:97–173

    PubMed  Google Scholar 

  5. Bhandari B, Naik MS, Nicholas DJD (1984) ATP production coupled to the denitrification of nitrate in Rhizobium japonicum, grown in cultures and in bacteroids from Glycine max. FEBS Lett 168:321–326

    Article  CAS  Google Scholar 

  6. Bradford MM (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 72:248–254

    Article  CAS  PubMed  Google Scholar 

  7. Bryan JK (1977) Molecular weights of protein multimers from polyacrylamide gel electrophoresis. Anal Biochem 78:513–519

    Article  CAS  PubMed  Google Scholar 

  8. Davis BJ (1964) Disc electrophoresis. II. Method and application to human serum proteins. Ann NY Acad Sci 121:404–436

    CAS  PubMed  Google Scholar 

  9. Delgado MJ, Bonnard N, Tresierra-Ayala A, Bedmar EJ, Muller P (2003) The Bradyrhizobium japonicum napEDABC genes encoding the periplasmic nitrate reductase are essential for nitrate respiration. Microbiology 149:3395–3403

    Article  CAS  PubMed  Google Scholar 

  10. Delgado MJ, Fernández-López M, Bedmar EJ (1998) Soluble and membrane-bound nitrate reductase from Bradyrhizobium japonicum bacteroids. Plant Physiol Biochem 36:279–283

    Article  CAS  Google Scholar 

  11. Fernández-López M, Olivares J, Bedmar EJ (1994) Two differentially regulated nitrate reductases required for nitrate-dependent, microaerobic growth of Bradyrhizobium japonicum. Arch Microbiol 162:310–315

    Google Scholar 

  12. Fernández-López M, Olivares J, Bedmar EJ (1996) Purification and characterization of the membrane-bound nitrate reductase isoenzymes of Bradyrhizobium japonicum. FEBS Lett 392:1–5

    PubMed  Google Scholar 

  13. Galibert F, Finan TM, Long SR, et al. (2001) The composite genome of the legume symbiont Sinorhizobium meliloti. Science 293:668–672

    CAS  PubMed  Google Scholar 

  14. Kaneko T, Nakamura Y, Sato S, et al. (2002) Complete genomic sequence of nitrogen-fixing symbiotic bacterium Bradyrhizobium japonicum USDA 110. DNA Res 9:189–197

    PubMed  Google Scholar 

  15. Kengen SW, Rikken GB, Hagen WR, van Ginkel CG, Stams AJ (1999) Purification and characterization of (per)chlorate reductase from the chlorate-respiring strain GR-1. J Bacteriol 181:6706–6711

    CAS  PubMed  Google Scholar 

  16. Kennedy IR, Riguard J, Trinchant JC (1975) Nitrate reductase from bacteroids of Rhizobium japonicum: enzyme characteristic and possible interaction with nitrogen fixation. Biochim Biophys Acta 397:24–35

    CAS  PubMed  Google Scholar 

  17. Kleinhofs A, Narayanan KR, Somers DA, Kuo TM, Warner RL (1986) Immunochemical methods for higher plant nitrate reductase. In: Linskens HF, Jackson JF (eds). Modern methods of plant analysis, new series, vol. 4. Berlin: Springer-Verlag, pp 190–211

    Google Scholar 

  18. Laemmli UK (1970) Cleavage of structural proteins during assembly of the head of bacteriophage T4. Nature 277:680–685

    Google Scholar 

  19. Moreno-Vivián C, Ferguson SJ (1998) Definition and distinction between assimilatory, dissimilatory and respiratory pathways. Mol Microbiol 29:664–666

    PubMed  Google Scholar 

  20. Moreno-Vivián C, Cabello P, Martinez-Luque M, Blasco R, Castillo F (1999) Prokaryotic nitrate reduction: molecular properties and functional distinction among bacterial nitrate reductases. J Bacteriol 181:6573–6584

    PubMed  Google Scholar 

  21. Philippot L, Højberg O (1999) Dissimilatory nitrate reductases in bacteria. Biochim Biophys Acta 1446:1–23

    CAS  PubMed  Google Scholar 

  22. Philippot L (2002) Denitrifying genes in bacterial and Archaeal genomes. Biochim Biophys Acta 1577:355–376

    CAS  PubMed  Google Scholar 

  23. Polcyn W, Luciński R (2001) Functional similarities of nitrate reduction from yellow lupine bacteroids to bacterial denitrification system. J Plant Physiol 158:829–834

    Article  CAS  Google Scholar 

  24. Polcyn W, Luciński R (2003) Aerobic and anaerobic nitrate and nitrite reduction in free-living cells of Bradyrhizobium sp. (Lupinus). FEMS Microbiol Lett 226:331–337

    Article  CAS  PubMed  Google Scholar 

  25. Richardson DJ, Berks BC, Russell DA, Spiro S, Taylor CJ (2001) Functional, biochemical and genetic diversity of prokaryotic nitrate reductases. Cell Mol Life Sci 58:165–178

    Article  CAS  PubMed  Google Scholar 

  26. Weiss RL (1976) Protoplast formation in Escherichia coli. J Bacteriol 128:668–670

    CAS  PubMed  Google Scholar 

  27. Zumft W (1997) Cell biology and molecular basis of denitrification. Microbiol Mol Biol Rev 61:533–616

    CAS  PubMed  Google Scholar 

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Acknowledgments

This work was supported by grants from the Polish State Committee for Scientific Research: 6 P04C 026 20 to R. L. and 2 P06R 077 27 to W. P. (years 2004–2007).

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  1. Department of Plant Physiology, Faculty of Biology, A. Mickiewicz University, Al. Niepodległości 14, 61-713, Poznań, Poland

    Władysław Polcyn & Robert Luciński

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  1. Władysław Polcyn
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  2. Robert Luciński
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Correspondence to Władysław Polcyn.

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Polcyn, W., Luciński, R. Dissimilatory Nitrate Reductase from Bradyrhizobium sp. (Lupinus): Subcellular Location, Catalytic Properties, and Characterization of the Active Enzyme Forms. Curr Microbiol 52, 231–237 (2006). https://doi.org/10.1007/s00284-005-0265-x

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  • DOI: https://doi.org/10.1007/s00284-005-0265-x

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